GSM3931693: THLE-2 EZH2 1st [ChIP-seq]; Homo sapiens; ChIP-Seq
Sample information curated by ChIP-Atlas
Antigen
Antigen Class
TFs and others
Antigen
EZH2
Cell type
Cell type Class
Liver
Cell type
THLE-2
Primary Tissue
Liver
Tissue Diagnosis
Normal
Attributes by original data submitter
Sample
source_name
cell line
cell line
THLE-2
cell type
normal liver epithelial cells
chip antibody
EZH2
Sequenced DNA Library
library_strategy
ChIP-Seq
library_source
GENOMIC
library_selection
ChIP
library_construction_protocol
The cells lysates was sonicated to generate DNA fragments of 200-500 bp, centrifuged for 10 min at 12,000 g, and the supernatant was directly used for immunoprecipitation. Place the tube containing the Pierce™ ChIP-grade Protein A/G Magnetic Beads on the magnetic, then, discard the supernatant. Wash the beads for three times with 1ml 0.1M sodium phosphate Buffer. Resuspend the beads with 200ul 0.1M sodium phosphate Buffer. Add 10 ug target antibody (CST, 5646s) and Rabbit IgG (CST, 2729s) antibody to the tube containing beads. Incubate mixture on a rotating wheel 2h at 4°C. Place the tube containing the beads on the magnetic stand for 1 minute. Then, discard the supernatant. Resuspend beads with the cells lysates. Rotate the sample for overnight at 4 °C. Place on the magnetic, then, discard the supernatant. The beads were sequentially washed with LiCl IP Wash Buffer (100 mM Tris (PH 7.5), 500 mM LiCl, 1% NP-40 and 0.5 deoxycholate) for five times, TE buffer (10Mm Tris (PH 8.0), 1mM EDTA (PH 8.0)) for one time, respectively. Resuspend sample with 100ul Elution Buffer (100 mM NaHCO3, 1% SDS) and reverse cross-linked by overnight incubation at 65 ℃. After sequential RNase A and proteinase K treatment, DNA fragments were purified by phenol extraction and ethanol precipitation. Resuspend the RNA pellet in10ul water. To generate libraries with the ThruPLEX® DNA-seq Kit (R400427, Rubicon Genomics) according the manufacturer's procedure. Briefly, purified DNA fragments were end-repaired, adenylated, ligated to adaptors and PCR amplification. Used the ThruPLEX® DNA-seq Kit. For high-throughput sequencing, the libraries were prepared following the manufacturer's instructions and applied to Illumina NextSeq 500 system for 100 nt pair-end sequencing by ABlife, Inc (Wuhan, China).